We have discovered a preparation of spontaneously beating cells from adult mammalian myocardium which are devoid of any measureable membrane potential. Electron microscopy of beating cells indicates no obvious structural membrane pathology. Preliminary experiments indicate sarcolemmal premeability to mono- and divalent cations, and suggest that contraction is initiated when a threshold (Ca)i is attained by diffusion, releasing additional Ca from sarcoplasmic reticulum (SR). Relaxation appears to be dependent on resequestration of Ca by SR. We propose to test this theory in several ways. We will measure contraction frequency and active tension of isolated cells while modifying the medium. If the above theory is correct, then contraction frequency and dp/dt, but not necessarily peak tension, should increase as a function of calcium content in the medium. Responsiveness to drugs should reflect mitochondrial and SR control, not sarcolemmal. Contractile performance should not vary with the Na/K in the medium. In addition, the cells should show measureable uptake of substances to which "intact" cells are impermeable, such as mannitol and inulin. If these conditions pertain, then we can conclude that regenerative release of calcium is the most likely mechanism controlling contraction in these cells. By implication, this will support the regenerative release theory which has been proposed for intact muscle. Furthermore, it will mean that we have a model in which intracellular events can be studied without physically breaking cells and fractionating organelles. We would then hope to use this model to study the effects of catecholamines and cyclic nucleotides on sarcoplasmic reticular, mitochondrial, and contractile protein metabolism and interaction.